title: Development and validation of an LC-ESI/MS/MS method with precolumn derivatization for the determination of betulin in rat plasma.
abstract: Neutral pentacyclic triterpenes with only one or two hydroxyl groups, such as betulin, are not easily ionized by electrospray ionization (ESI). However, because betulin is reactive and neutral, derivatization may improve ionization efficiency. In the present study, the potency of different derivatization reagents was evaluated and p-toluenesulfonyl isocyanate (PTSI) was proven to be the optimal. The derivative generated by the reaction of betulin with PTSI was ionizable and fragmentable in the negative mode by liquid chromatography-electrospray ionization/tandem mass spectrometry (LC-ESI/MS/MS). Based on this chemical derivatization, an LC-ESI/MS/MS method was developed and validated for the quantification of betulin in rat plasma. The sample was extracted with ethyl acetate, derivatized with PTSI, separated on an ACQ UPLC BEH phenyl column, and analyzed in negative multiple reaction monitoring (MRM) mode. The calibration curve was linear over the betulin concentration range 2.5-200ng/mL. The lower limit of quantification was 2.5ng/mL. The inter- and intra-day accuracy and precision were within ±15%. Betulin recoveries were 86.7% or higher at three quality control levels (5, 50, and 160ng/mL). This validated method was subsequently applied to a pharmacokinetic study of betulin in rat plasma after oral administration.
authors.affiliation: Alkali Soil Natural Environmental Science Center, Northeast Forestry University, Key Laboratory of Saline-Alkali Vegetation Ecology Restoration in Oil Field, Ministry of Education, Harbin 150040, China. Electronic address: huzhiwei.haerbin@yahoo.com.cn.
authors.forename: Zhiwei
authors.initials: Z
authors.lastname: Hu
authors.forename: Na
authors.initials: N
authors.lastname: Guo
authors.forename: Ziming
authors.initials: Z
authors.lastname: Wang
authors.forename: Yong
authors.initials: Y
authors.lastname: Liu
authors.forename: Yu
authors.initials: Y
authors.lastname: Wang
authors.forename: Weimin
authors.initials: W
authors.lastname: Ding
authors.forename: Dehui
authors.initials: D
authors.lastname: Zhang
authors.forename: Yang
authors.initials: Y
authors.lastname: Wang
authors.forename: Xiufeng
authors.initials: X
authors.lastname: Yan
mesh_terms: D000818:Animals; D002853:Chromatography, Liquid / Q000379:methods*; D016014:Linear Models; D008297:Male; D051381:Rats; D017208:Rats, Wistar; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D021241:Spectrometry, Mass, Electrospray Ionization; D053719:Tandem Mass Spectrometry / Q000379:methods*; D014105:Tosyl Compounds; D014315:Triterpenes / Q000097:blood* / Q000737:chemistry / Q000493:pharmacokinetics; D000094062:Betulinic Acid
chemical_list: D014105:Tosyl Compounds; D014315:Triterpenes; C002503:betulin; D000094062:Betulinic Acid
keywords: Betulin; LC–ESI/MS/MS; Pharmacokinetics; Precolumn derivatization; p-Toluenesulfonyl isocyanate